DIFFERENT MECHANISMS CONTROL SIGNAL-INDUCED DEGRADATION AND BASAL TURNOVER OF THE NF-KAPPA-B INHIBITOR I-KAPPA-B-ALPHA IN-VIVO

The transcription factor NF-kappa B is sequestered in the cytoplasm by a family of I kappa B molecules. Upon cellular stimulation with diver se agents, one of these molecules, I kappa B alpha, is rapidly phospho rylated and subsequently degraded. This process triggers nuclear trans location of NF-kappa B and the successive activation of target genes. Independent of its rapid stimulation-induced breakdown, I kappa B alph a is inherently unstable and undergoes a continuous turnover. To compa re the mechanisms and protein domains involved in inducible and basal degradation of I kappa B alpha in intact cells we employed a transfect ion strategy using tagged I kappa B alpha and ubiquitin molecules. We show that tumor necrosis factor alpha (TNF alpha) induced breakdown of I kappa B alpha but not its basal turnover coincides with ubiquitinat ion in the aminoterminal signal response domain (SRD) of I kappa B alp ha. Neither the SRD nor the carboxy-terminal PEST sequence is needed f or basal turnover, which instead depends only on the core ankyrin repe at domain. Despite the differences in the requirements of protein doma ins and ubiquitin-conjugation for both degradation pathways, each one is mediated by the proteasome. This finding is important for understan ding alternative modes of controlling NF-kappa B activity.